The present invention relates generally to the field of oncology, and to methods and pharmaceutical compositions for enhancing the activity of a cancer chemotherapeutic agent. More particularly, the present invention concerns the use of a 3-aryloxy-3-phenylpropylamine such as fluoxetine [(N-methyl 3-(p-trifluoromethylphenoxy)-3-phenylpropylamine] as a chemosensitizer for enhancing the cytotoxicity of a chemotherapeutic agent, especially in drug-resistant tumors and more particularly in the case of Multidrug Resistance (MDR). Methods and compositions are provided for the treatment of cancers such as, but not limited to, leukemia, lymphoma, carcinoma and sarcoma (including glioma) using a 3-aryloxy-3-phenylpropylamine, fluoxetine in particular, as a chemosensitizer.
Many of the most prevalent forms of human cancer resist effective chemotherapeutic intervention. Some tumor populations, especially adrenal, colon, jejunal, kidney and liver carcinomas, appear to have drug-resistant cells at the outset of treatment (Barrows, L. R., xe2x80x9cAntineoplastic and Immunoactive Drugsxe2x80x9d, Chapter 75, pp 1236-1262, in: Remington: The Science and Practice of Pharmacy, Mack Publishing Co. Easton, Pa., 1995). In other cases, a resistance-conferring genetic change occurs during treatment; the resistant daughter cells then proliferate in the environment of the drug. Whatever the cause, resistance often terminates the usefulness of an antineoplastic drug.
Clinical studies suggest that a common form of multidrug resistance in human cancers results from the expression of the MDR1 gene that encodes P-glycoprotein. This glycoprotein functions as a plasma membrane, energy-dependent, multidrug efflux pump that reduces the intracellular concentration of cytotoxic drugs. This mechanism of resistance may account for de novo resistance in common tumors, such as colon cancer and renal cancer, and for acquired resistance, as observed in common hematologic tumors such as acute nonlymphocytic leukemia and malignant lymphomas. Although this type of drug resistance may be common, it is by no means the only mechanism by which cells become drug resistant. MDR is effected via an extrusion mechanism (Tan B, Piwnica-Worms D, Ratner L., Multidrug resistance transporters and modulation. Curr. Opin. Oncol, September 2000;12(5):450-8). The influx of chemotherapeutic drugs into cells is mainly by passive diffusion across the cell membrane, driven by the drug""s electrochemical-potential gradient. In MDR cells there are energy-dependant extrusion channels that actively pump the drug out of the cells, reducing it""s intracellular concentration below lethal threshold. The first pump identified was named Pgp (for P-glycoprotein), the second was named MRP (for Multidrug Resistant associate Protein) and several more have been identified in recent years (Tan et al. 2000, ibid.). All of them are naturally occurring proteins, and their physiological roles are assumed to involve detoxification of cells. In MDR cells they are present, for reasons yet unknown, in a significantly higher number of copies than in other non-MDR cells.
Chemical modification of cancer treatment involves the use of agents or maneuvers that are not cytotoxic in themselves, but modify the host or tumor so as to enhance anticancer therapy. Such agents are called chemosensitizers. Pilot studies using chemosensitizers indicate that these agents may reverse resistance in a subset of patients. These same preliminary studies also indicate that drug resistance is multifactorial, because not all drug-resistant patients have P-glycoprotein-positive tumor cells and only a few patients appear to benefit from the use of current chemosensitizers.
Chemosensitization research has centered on agents that reverse or modulate multidrug resistance in solid tumors (MDR1, P-glycoprotein). Chemosensitizers known to modulate P-glycoprotein function include: calcium channel blockers (verapamil, indicated for the treatment of hypertension), calmodulin inhibitors (trifluoperazine), indole alkaloids (reserpine), quinolines (quinine), lysosomotropic agents (chloroquine), steroids, (progesterone), triparanol analogs (tamoxifen), detergents (cremophor EL), and cyclic peptide antibiotics (cyclosporines, indicated to prevent host vs. graft disease) (DeVita, V. T., et al., in Cancer, Principles and Practice of Oncology, 4th ed., J. B. Lippincott Co., Philadelphia, Pa., pp 2661-2664, 1993; Sonneveld P, Wiemer E. Inhibitors of multidrug resistance., Curr Opin Oncol November 1997;9(6):543-8).
A review of studies where chemosensitizing agents were used concluded the following: (i) cardiovascular side effects associated with continuous, high-dose intravenous verapamil therapy are significant and dose-limiting; (ii) dose-limiting toxicities of the chemosensitizers, trifluoperazine and tamoxifen, was attributed to the inherent toxicity of the chemosensitizer and not due to enhanced chemotherapy toxicity; (iii) studies using high doses of cyclosporin A as a chemosensitizer found hyperbilirubinemia as a side effect; and (iv) further research is clearly needed to develop less toxic and more efficacious chemosensitizers to be used clinically (DeVita et al., 1993, ibid.).
For example, while verapamil is effective in hypertension treatment at the 2-4 xcexcM range, for MDR reversal it requires the dose range of 10-15 xcexcM, while at 6 xcexcM it is already in the toxic domain.
Tumors that are considered drug-sensitive at diagnosis but acquire an MDR phenotype at relapse, pose an especially difficult clinical problem. At diagnosis, only a minority of tumor cells may express P-glycoprotein and treatment with chemotherapy provides a selection advantage for the few cells that are P-glycoprotein positive early in the course of disease. Another possibility is that natural-product-derived chemotherapy actually induces the expression of MDR1, leading to P-glycoprotein-positive tumors at relapse. Using chemosensitizers early in the course of disease may prevent the emergence of MDR by eliminating the few cells that are P-glycoprotein positive at the beginning. In vitro studies have shown that selection of drug-resistant cells by combining verapamil and doxorubicin does prevent the emergence of P-glycoprotein, but that an alternative drug resistance mechanism develops, which is secondary to altered topoisomerase II function (Dalton, W. S., Proc. Am. Assoc. Cancer Res. 31:520, 1990).
More efficacious and less toxic chemosensitizers are urgently needed to improve the outcome of chemotherapy. Clinical utility of a chemosensitizer depends upon its ability to enhance the cytotoxicity of a chemotherapeutic drug and also on its low toxicity in vivo. The present inventors have addressed these problems and provide herein a new class of chemosensitizers that permit new approaches in cancer treatment.
3-Aryloxy-3-phenylpropylamines and their use to treat depression are described in, for example, U.S. Pat. Nos. 4,018,895 and 6,258,853. Fluoxetine [(N-methyl 3-(p-trifluoromethylphenoxy)-3-phenylpropylamine], known better by its commercial name Prozac, is a well-known approved drug, indicated for psychiatric treatments (Cookson J, Duffett R., Fluoxetine: therapeutic and undesirable effects. Hosp Med August 1998;59(8):622-6). It is known to be an SSRI (Selective Serotonin Reuptake Inhibition) agent, and this activity is considered to be related to its mechanism of action in its capacity as a psychiatric drug (Cookson et al., 1998, ibid.).
3-Aryloxy-3-phenylpropylamines in general and fluoxetine in particular have not hitherto indicated as chemosensitizers for the treatment of cancer.
While reducing the present invention to practice it was unexpectedly found that fluoxetine, a member of the 3-aryloxy-3-phenylpropylamines compounds, induces a significant enhancement of the cytotoxic effect of conventional chemotherapeutic drugs at a dose range well below its toxicity limits.
The present invention provides a method of chemosensitization comprising administering at least one chemotherapeutic agent and at least one 3-aryloxy-3-phenylpropylamine to a subject in need thereof. xe2x80x9cChemosensitizationxe2x80x9d, as used herein, means that a 3-aryloxy-3-phenylpropylamine increases or enhances the cytotoxicity of a chemotherapeutic agent compared to a level of cytotoxicity seen by that agent in the absence of 3-aryloxy-3-phenylpropylamine. That is, 3-aryloxy-3-phenylpropylamine xe2x80x9csensitizesxe2x80x9d a cancer cell to the effects of the chemotherapeutic agent, allowing the agent to be more effective. 3-Aryloxy-3-phenylpropylamines are not known, and are shown herein not to have anti-cancer chemotherapeutic activity on their own.
An embodiment of the present invention is a method of treating cancer in a subject comprising administering a chemotherapeutic agent and a 3-aryloxy-3-phenylpropylamine to the subject. The cancer may be leukemia, lymphoma, carcinoma, or sarcoma. In a preferred embodiment, a patient having a form of cancer for which chemotherapy is indicated is administered a dose of 3-aryloxy-3-phenylpropylamine at intervals with each dose of the chemotherapeutic agent.
In another aspect of the invention, 3-aryloxy-3-phenylpropylamines may be used as a topical chemosensitizer. Table 1 below indicates that 5-fluorouracil, for example, is used topically for premalignant skin lesions. The inventors envision the use of 3-aryloxy-3-phenylpropylamines to enhance the cytotoxicity of topical chemotherapeutic agents.
A method for selecting a chemotherapeutic agent for which 3-aryloxy-3-phenylpropylamine is a chemosensitizer is a further embodiment of the present invention. The method comprises (i) assaying cytotoxicity of a candidate chemotherapeutic agent in the presence and in the absence of a 3-aryloxy-3-phenylpropylamine; and (ii) selecting a candidate chemotherapeutic agent as a chemotherapeutic agent for which 3-aryloxy-3-phenylpropylamine is a chemosensitizer when the cytotoxicity of the candidate agent is greater in the presence of 3-aryloxy-3-phenylpropylamine than in the absence of 3-aryloxy-3-phenylpropylamine. A presently preferred in vitro assay is the MTT cytotoxicity assay cited in the examples section. An exemplary in vivo assay is described in, for example, U.S. Pat. No. 5,776,925, which is incorporated herein by reference.
A 3-aryloxy-3-phenylpropylamine used as a chemosensitiszer in accordance with the teachings of the present invention is preferably of the formula: 
wherein each Rxe2x80x2 is independently hydrogen or methyl;
R is naphthyl or 
Rxe2x80x3 and Rxe2x80x2xe2x80x3 are halo, trifluoromethyl, C1-C4 alkyl, C1-C3 alkoxy or C3-C4 alkenyl; and
n and m are 0, 1 or 2; and acid addition salts thereof formed with pharmaceutically acceptable acids.
In the above formula when R is naphthyl, it can be either alpha-naphthyl or beta-naphthyl. Rxe2x80x3 and Rxe2x80x2xe2x80x3 when they are halo, C1-C4 alkyl, C1-C3 alkyloxy or C3-C4 alkenyl represent, illustratively, the following atoms or groups: fluoro, chloro, bromo, iodo, methyl, ethyl, isopropyl, n-propyl, n-butyl, isobutyl, sec-butyl, t-butyl, methoxy, ethoxy, n-propoxy, isopropoxy, allyl, methallyl, crotyl and the like. R thus can represent o, m and p-trifluoromethylphenyl, o, m and p-chlorophenyl, o, m and p-bromophenyl, o, m and p-fluorophenyl, o, m and p-tolyl, xylyl including all position isomers, o, m and p-anisyl, o, m and p-allylphenyl, o, m and p-methylallylphenyl, o, m and p-phenetolyl(ethoxyphenyl), 2,4-dichlorophenyl, 3,5-difluorophenyl, 2-methoxy-4-chlorophenyl, 2-methyl-4-chlorophenyl, 2-ethyl-4-bromophenyl, 2,4,6-trimethylphenyl, 2-fluoro-4-trifluoromethylphenyl, 2,4,6-trichlorophenyl, 2,4,5-trichlorophenyl and the like.
Also included within the scope of this invention are the pharmaceutically-acceptable salts of the amine bases represented by the above formula formed with non-toxic acids. These acid addition salts include salts derived from inorganic acids such as: hydrochloric acid, nitric acid, phosphoric acid, sulfuric acid, hydrobromic acid, hydriodic acid, nitrous acid, phosphorous acid and the like, as well as salts of non-toxic organic acids including aliphatic mono and dicarboxylates, phenyl-substituted alkanoates, hydroxy alkanoates and alkanedioates, aromatic acids, aliphatic and aromatic sulfonic acids etc. Such pharmaceutically-acceptable salts thus include: sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, nitrate, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate, chloride, bromide, iodide, fluorodide, acetate, propionate, decanoate, caprylate, acrylate, formate, isobutyrate, caprate, heptanoate, propiolate, oxalate, malonate, succinate, suberate, sebacate, fumarate, maleate, butyne-1,4-dioate, hexyne-1,6-dioate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, phthalate, terephthalate, benzenesulfonates, toluenesulfonate, chlorobenzenesulfonate, xylenesulfonate, phenylacetate, phenylpropionate, phenylbutyrate, citrate, lactate, beta-hydroxybutyrate, glycollate, malate, tartrate, methanesulfonate, propanesulfonates, naphthalene-1-sulfonate, naphthalene-2-sulfonate, mandelate and the like salts.
Compounds illustrative of the scope of this invention include the following:
3-(p-isopropoxyphenxoy)-3-phenylpropylamine methanesulfonate;
N,N-dimethyl 3-(3xe2x80x2,4xe2x80x2-dimethoxyphenoxy)-3-phenylpropylamine p-hydroxybenzoate;
N,N-dimethyl 3-(alpha-naphthoxy)-3-phenylpropylamine bromide;
N,N-dimethyl 3-(beta-naphthoxy)-3-phenyl-1-methylpropylamine iodide;
3-(2xe2x80x2-methyl-4xe2x80x2,5xe2x80x2-dichlorophenoxy)-3-phenylpropylamine nitrate;
3-(p-t-butylphenoxy)-3-phenylpropylamine glutarate;
N-methyl 3-(2xe2x80x2-chloro-p-tolyloxy)-3-phenyl-1-methylpropylamine lactate;
3-(2xe2x80x2,4xe2x80x2-dichlorophenoxy)-3-phenyl-2-methylpropylamine citrate;
N,N-dimethyl 3-(m-anisyloxy)-3-phenyl-1-methylpropylamine maleate;
N-methyl 3-(p-tolyloxy)-3-phenylpropylamine sulfate;
N,N-dimethyl 3-(2xe2x80x2,4xe2x80x2-difluorophenoxy)-3-phenylpropylamine 2,4-dinitrobenzoate;
3-(o-ethylphenoxy)-3-phenylpropylamine dihydrogen phosphate;
N-methyl-(2xe2x80x2-chloro-4xe2x80x2-isopropylphenoxy)-3-phenyl-2-methylpropylamine maleate;
N,N-dimethyl 3-(2xe2x80x2-alkyl-4xe2x80x2-fluorophenoxy)-3-phenylpropylamine succinate;
N,N-dimethyl 3-(o-isopropoxyphenoxy)-3-phenyl-propylamine phenylacetate;
N,N-dimethyl 3-(o-)bromophenoxy)-3-phenyl-propylamine beta-phenylpropionate;
N-methyl 3-(p-iodophenoxy)-3-phenyl-propylamine propiolate;
N-methyl 3-(3-n-propylphenoxy)-3-phenyl-propylamine decanoate; and preferably,
N-methyl 3-(p-trifluoromethylphenoxy)-3-phenylpropylamine.
The present invention successfully addresses the shortcomings of the presently known configurations by identifying a new chemosensitizer which efficiently act at concentrations well below its toxicity and which is of particular efficacy in chemosensitizing multi drug resistant (MDR) cells.